Current perspectives on mesenchymal stem cells as a potential therapeutic strategy for non-alcoholic fatty liver disease

Non-alcoholic fatty liver disease(NAFLD)has emerged as a significant health challenge,characterized by its widespread prevalence,intricate natural progression and multifaceted pathogenesis.Although NAFLD initially presents as benign fat accumulation,it may progress to steatosis,non-alcoholic steatohepatitis,cirrhosis,and hepatocellular carcinoma.Mesenchymal stem cells(MSCs)are recognized for their intrinsic self-renewal,superior biocompatibility,and minimal immunogenicity,positioning them as a therapeutic innovation for liver diseases.Therefore,this review aims to elucidate the potential roles of MSCs in alleviating the progression of NAFLD by alteration of underlying molecular pathways,including glycolipid metabolism,inflammation,oxidative stress,endoplasmic reticulum stress,and fibrosis.The insights are expected to provide further understanding of the potential of MSCs in NAFLD therapeutics,and support the development of MSC-based therapy in the treatment of NAFLD.

Three-dimensional cell-based strategies for liver regeneration

Liver regeneration and the development of effective therapies for liver failure remain formidable challenges in modern medicine.In recent years,the utilization of 3D cell-based strategies has emerged as a promising approach for addressing these urgent clinical requirements.This review provides a thorough analysis of the application of 3D cell-based approaches to liver regeneration and their potential impact on patients with end-stage liver failure.Here,we discuss various 3D culture models that incorporate hepatocytes and stem cells to restore liver function and ameliorate the consequences of liver failure.Furthermore,we explored the challenges in transitioning these innovative strategies from preclinical studies to clinical applications.The collective insights presented herein highlight the significance of 3D cell-based strategies as a transformative paradigm for liver regeneration and improved patient care.

Comprehensive Understanding of Immune Cells in The Pathogenesis of Non-alcoholic Fatty Liver Disease

Non-alcoholic fatty liver disease(NAFLD)is the most common chronic liver disease,defined by several phases,ranging from benign fat accumulation to non-alcoholic steatohepatitis(NASH),which can lead to liver cancer and cirrhosis.Although NAFLD is a disease of disordered metabolism,it also involves several immune cell-mediated inflammatory processes,either promoting and/or suppressing hepatocyte inflammation through the secretion of pro-inflammatory and/or anti-inflammatory factors to influence the NAFLD process.However,the underlying disease mechanism and the role of immune cells in NAFLD are still under investigation,leaving many open-ended questions.In this review,we presented the recent concepts about the interplay of immune cells in the onset and pathogenesis of NAFLD.We also highlighted the specific non-immune cells exhibiting immunological properties of therapeutic significance in NAFLD.We hope that this review will help guide the development of future NAFLD therapeutics.

Innovative mesenchymal stem cell treatments for fatty liver disease

The incidence of non-alcoholic fatty liver disease(NAFLD)and alcohol-associated liver disease(ALD)is increasing year by year due to changes in the contemporary environment and dietary structure,and is an important public health problem worldwide.There is an urgent need to continuously improve the understanding of their disease mechanisms and develop novel therapeutic strategies.Mesenchymal stem cells(MSCs)have shown promise as a potential therapeutic strategy in therapeutic studies of NAFLD and ALD.NAFLD and ALD have different triggers and their specific mechanisms of disease progression are different,but both involve disease processes such as hepatocellular steatosis and potential fibrosis,cirrhosis,and even hepatocellular carcinoma.MSCs have metabolic regulatory,anti-apoptotic,antioxidant,and immunomodulatory effects that together promote liver injury repair and functional recovery,and have demonstrated positive results in preclinical studies.This editorial is a continuum of Jiang et al’s review focusing on the advantages and limitations of MSCs and their derivatives as therapeutics for NAFLD and ALD.They detail how MSCs attenuate the progression of NAFLD by modulating molecular pathways involved in glucolipid metabolism,inflammation,oxidative stress,endoplasmic reticulum stress,and fibrosis.Based on recent advances,we discuss MSCs and their derivatives as therapeutic strategies for NAFLD and ALD,providing useful information for their clinical application.

Mechanism of mesenchymal stem cells in liver regeneration:Insights and future directions

Mesenchymal stem cells(MSCs)are a prevalent source for stem cell therapy and play a crucial role in modulating both innate and adaptive immune responses.Non-alcoholic fatty liver disease(NAFLD)is characterized by the accumulation of triglycerides in liver cells and involves immune system activation,leading to histological changes,tissue damage,and clinical symptoms.A recent publication by Jiang et al,highlighted the potential of MSCs to mitigate in NAFLD progression by targeting various molecular pathways,including glycolipid metabolism,inflammation,oxidative stress,endoplasmic reticulum stress,and fibrosis.In this editorial,we comment on their research and discuss the efficacy of MSC therapy in treating NAFLD.

Cellular strategies to induce immune tolerance after liver transplantation:Clinical perspectives

Liver transplantation(LT)has become the most efficient treatment for pediatric and adult end-stage liver disease and the survival time after transplantation is becoming longer due to the development of surgical techniques and perioperative management.However,long-term side-effects of immunosuppressants,like infection,metabolic disorders and malignant tumor are gaining more attention.Immune tolerance is the status in which LT recipients no longer need to take any immunosuppressants,but the liver function and intrahepatic histology maintain normal.The approaches to achieve immune tolerance after transplantation include spontaneous,operational and induced tolerance.The first two means require no specific intervention but withdrawing immunosuppressant gradually during follow-up.No clinical factors or biomarkers so far could accurately predict who are suitable for immunosuppressant withdraw after transplantation.With the understanding to the underlying mechanisms of immune tolerance,many strategies have been developed to induce tolerance in LT recipients.Cellular strategy is one of the most promising methods for immune tolerance induction,including chimerism induced by hematopoietic stem cells and adoptive transfer of regulatory immune cells.The safety and efficacy of various cell products have been evaluated by prospective preclinical and clinical trials,while obstacles still exist before translating into clinical practice.Here,we will summarize the latest perspectives and concerns on the clinical application of cellular strategies in LT recipients.

Mesenchymal stem cells: A promising therapeutic avenue for nonalcoholic fatty liver disease

Non-alcoholic fatty liver disease(NAFLD)is a pressing global health concern that is associated with metabolic syndrome and obesity.On the basis of the insights provided by Jiang et al,this editorial presents an exploration of the potential of mesenchymal stem cells(MSCs)for NAFLD treatment.MSCs have numerous desirable characteristics,including immunomodulation,anti-inflammatory pro-perties,and tissue regeneration promotion,rendering them attractive candidates for NAFLD treatment.Recent preclinical and early clinical studies have high-lighted the efficacy of MSCs in improving liver function and reducing disease severity in NAFLD models.However,MSC heterogeneity,long-term safety concerns,and unoptimized therapeutic protocols remain substantial challenges.Addressing these challenges through standardized protocols and rigorous clinical trials is essential to the safe and successful application of MSCs in NAFLD mana-gement.Continued research into MSC mechanisms and therapeutic optimization is required to improve treatments for NAFLD and related liver diseases.

Perilipin 5 regulates hepatic stellate cell activation and high-fat diet-induced non-alcoholic fatty liver disease

Background:Nonalcoholic fatty liver disease(NAFLD)is one of the most common chronic liver diseases globally.Hepatic stellate cells(HSCs)are the major effector cells of liver fibrosis.HSCs contain abundant lipid droplets(LDs)in their cytoplasm during quiescence.Perilipin 5(PLIN 5)is a LD surface-associated protein that plays a crucial role in lipid homeostasis.However,little is known about the role of PLIN 5 in HSC activation.Methods:PLIN 5 was overexpressed in HSCs of Sprague–Dawley rats by lentivirus transfection.At the same time,PLIN 5 gene knockout mice were constructed and fed with a high-fat diet(HFD)for 20 weeks to study the role of PLIN 5 in NAFLD.The corresponding reagent kits were used to measure TG,GSH,Caspase 3 activity,ATP level,and mitochondrial DNA copy number.Metabolomic analysis of mice liver tissue metabolism was performed based on UPLC-MS/MS.AMPK,mitochondrial function,cell proliferation,and apoptosis-related genes and proteins were detected by western blotting and qPCR.Results:Overexpression of PLIN 5 in activated HSCs led to a decrease in ATP levels in mitochondria,inhibition of cell proliferation,and a significant increase in cell apoptosis through AMPK activation.In addition,compared with the HFD-fed C57BL/6J mice,PLIN 5 knockout mice fed with HFD showed reduced liver fat deposition,decreased LD abundance and size,and reduced liver fibrosis.Conclusion:These findings highlight the unique regulatory role of PLIN 5 in HSCs and the role of PLIN 5 in the fibrosis process of NAFLD.

Ginsenoside Rb1 induces hepatic stellate cell ferroptosis to alleviate liver fibrosis via the BECN1/SLC7A11 axis

Liver fibrosis is primarily driven by the activation of hepatic stellate cells(HSCs),a process associated with ferroptosis.Ginsenoside Rb1(GRb1),a major active component extracted from Panax ginseng,inhibits HSC activation.However,the potential role of GRb1 in mediating HSC ferroptosis remains unclear.This study examined the effect of GRb1 on liver fibrosis both in vivo and in vitro,using CCl4-induced liver fibrosis mouse model and primary HSCs,LX-2 cells.The findings revealed that GRb1 effectively inactivated HSCs in vitro,reducing alpha-smooth muscle actin(a-SMA)and type I collagen(Col1A1)levels.Moreover,GRb1 significantly alleviated CCl4-induced liver fibrosis in vivo.From a mechanistic standpoint,the ferroptosis pathway appeared to be central to the antifibrotic effects of GRb1.Specifically,GRb1 promoted HSC ferroptosis both in vivo and in vitro,characterized by increased glutathione depletion,malondialdehyde production,iron overload,and accumulation of reactive oxygen species(ROS).Intriguingly,GRb1 increased Beclin 1(BECN1)levels and decreased the System Xc-key subunit SLC7A11.Further experiments showed that BECN1 silencing inhibited GRb1-induced effects on HSC ferroptosis and mitigated the reduction of SLC7A11 caused by GRb1.Moreover,BECN1 could directly interact with SLC7A11,initiating HSC ferroptosis.In conclusion,the suppression of BECN1 counteracted the effects of GRb1 on HSC inactivation both in vivo and in vitro.Overall,this study highlights the novel role of GRb1 in inducing HSC ferroptosis and promoting HSC inactivation,at least partly through its modulation of BECN1 and SLC7A11.

Enhancing the functionality of mesenchymal stem cells:An attractive treatment strategy for metabolic dysfunction-associated steatotic liver disease?

The intrinsic heterogeneity of metabolic dysfunction-associated fatty liver disease(MASLD)and the intricate pathogenesis have impeded the advancement and clinical implementation of therapeutic interventions,underscoring the critical demand for novel treatments.A recent publication by Li et al proposes mesenchymal stem cells as promising effectors for the treatment of MASLD.This editorial is a continuum of the article published by Jiang et al which focuses on the significance of strategies to enhance the functionality of mesenchymal stem cells to improve efficacy in curing MASLD,including physical pretreatment,drug or chemical pretreatment,pretreatment with bioactive substances,and genetic engineering.

Glycogen metabolism-mediated intercellular communication in the tumor microenvironment influences liver cancer prognosis

Glycogen metabolism plays a key role in the development of hepatoellular carcinoma(HCC),but the function of glycogen metabolism genes in the tumor microenvironment(TME)is still to be elucidated.Single cell RNA-seq data were obtained from ten HCC tumor samples totaling 64,545 cells and 65 glycogen metabolism genes were analyzed bya nonnegative matrix factorization(NMF).The prognosis and immune response of new glycogen TME cell dusters were predicted by using HCC and immunotherapy cohorts from public databases.HOC single cell analysis was divided into fibroblasts,NT T cells,macrophages,endothelial clls,and B cells,which were separately divided into new cell clusters by glycogen metabolism gene annotation.Pseudo temporal trajectory analysis demonstrated the temporal differentiation trajectory of different glycogen subtype cell dusters.Cellular communication analysis revealed extensive interactions between endothelial cells with glycogen metabolizing TME cell.related subtypes and diferent glycogen subtype cell clusters.SCENIC analysis of transcription factors upstream of TME cell clusters with different glycogen metabolism.In addition,TME cell dusters of glycogen metabolism were found to be enriched in expression in CAF subtypes,CD8 depleted,M1,and M2 types.Bulk seq analysis showed the prognostic signifcance of glycogen metabolism.mediated TME cell dusters in HCC,while a significant immune response was found in the immunotherapy cohort in patients treated with immune checkpoint blockade(ICB),especially for CAFs,T cells,and macrophages In summary,our study reveals for the first time that glycogen metabolism mediates intercellular communication in the hepatocellular carcinoma microenvironment while elucidating the anti-tumor mechanisms and immune prognostic responses of different subtypes of cell dusters.

Red cell distribution width/platelet ratio estimates the 3-year risk of decompensation in Metabolic Dysfunction-Associated Steatotic Liver Disease-induced cirrhosis

BACKGROUND For compensated advanced chronic liver disease(cACLD)patients,the first decompensation represents a dramatically worsening prognostic event.Based on the first decompensation event(DE),the transition to decompensated advanced chronic liver disease(dACLD)can occur through two modalities referred to as acute decompensation(AD)and non-AD(NAD),respectively.Clinically Significant Portal Hypertension(CSPH)is considered the strongest predictor of decompensation in these patients.However,due to its invasiveness and costs,CSPH is almost never evaluated in clinical practice.Therefore,recognizing noninvasively predicting tools still have more appeal across healthcare systems.The red cell distribution width to platelet ratio(RPR)has been reported to be an indicator of hepatic fibrosis in Metabolic Dysfunction-Associated Steatotic Liver Disease(MASLD).However,its predictive role for the decompensation has never been explored.AIM In this observational study,we investigated the clinical usage of RPR in predicting DEs in MASLD-related cACLD patients.METHODS Fourty controls and 150 MASLD-cACLD patients were consecutively enrolled and followed up(FUP)semiannually for 3 years.At baseline,biochemical,clinical,and Liver Stiffness Measurement(LSM),Child-Pugh(CP),Model for End-Stage Liver Disease(MELD),aspartate aminotransferase/platelet count ratio index(APRI),Fibrosis-4(FIB-4),Albumin-Bilirubin(ALBI),ALBI-FIB-4,and RPR were collected.During FUP,DEs(timing and modaities)were recorded.CSPH was assessed at the baseline and on DE occurrence according to the available Clinical Practice Guidelines.RESULTS Of 150 MASLD-related cACLD patients,43(28.6%)progressed to dACLD at a median time of 28.9 months(29 NAD and 14 AD).Baseline RPR values were significantly higher in cACLD in comparison to controls,as well as MELD,CP,APRI,FIB-4,ALBI,ALBI-FIB-4,and LSM in dACLD-progressing compared to cACLD individuals[all P<0.0001,except for FIB-4(P:0.007)and ALBI(P:0.011)].Receiving operator curve analysis revealed RPR>0.472 and>0.894 as the best cut-offs in the prediction respectively of 3-year first DE,as well as its superiority compared to the other non-invasive tools examined.RPR(P:0.02)and the presence of baseline-CSPH(P:0.04)were significantly and independently associated with the DE.Patients presenting baseline-CSPH and RPR>0.472 showed higher risk of decompensation(P:0.0023).CONCLUSION Altogether these findings suggest the RPR as a valid and potentially applicable non-invasive tool in the prediction of timing and modalities of decompensation in MASLD-related cACLD patients.

Blood cell counts and nonalcoholic fatty liver disease: Evidence from Mendelian randomization analysis

BACKGROUND Previous research has highlighted correlations between blood cell counts and chronic liver disease.Nonetheless,the causal relationships remain unknown.AIM To evaluate the causal effect of blood cell traits on liver enzymes and nonalcoholic fatty liver disease(NAFLD)risk.METHODS Independent genetic variants strongly associated with blood cell traits were extracted from a genome-wide association study(GWAS)conducted by the Blood Cell Consortium.Summary-level data for liver enzymes were obtained from the United Kingdom Biobank.NAFLD data were obtained from a GWAS meta-analysis(8434 cases and 770180 controls,discovery dataset)and the Fingen GWAS(2275 cases and 372727 controls,replication dataset).This analysis was conducted using the inverse-variance weighted method,followed by various sensitivity analyses.RESULTS One SD increase in the genetically predicted haemoglobin concentration(HGB)was associated with aβof 0.0078(95%CI:0.0059-0.0096),0.0108(95%CI:0.0080-0.0136),0.0361(95%CI:0.0156-0.0567),and 0.0083(95%CI:00046-0.0121)for alkaline phosphatase(ALP),alanine aminotransferase(ALT),aspartate aminotransferase,and gammaglutamyl transferase,respectively.Genetically predicted haematocrit was associated with ALP(β=0.0078,95%CI:0.0052-0.0104)and ALT(β=0.0057,95%CI:0.0039-0.0075).Genetically determined HGB and the reticulocyte fraction of red blood cells increased the risk of NAFLD[odds ratio(OR)=1.199,95%CI:1.087-1.322]and(OR=1.157,95%CI:1.071-1.250).The results of the sensitivity analyses remained significant.CONCLUSION Novel causal blood cell traits related to liver enzymes and NAFLD development were revealed through Mendelian randomization analysis,which may facilitate the diagnosis and prevention of NAFLD.

Subcellular distribution of prohibitin 1 in rat liver during liver regeneration and its cellular implication

BACKGROUND The function of prohibitin 1(Phb1)during liver regeneration(LR)remains relatively unexplored.Our previous research identified downregulation of Phb1 in rat liver mitochondria 24 h after 70%partial hepatectomy(PHx),as determined by subcellular proteomic analysis.AIM To investigate the potential role of Phb1 during LR.METHODS We examined changes in Phb1 mRNA and protein levels,subcellular distribution,and abundance in rat liver during LR following 70%PHx.We also evaluated mitochondrial changes and apoptosis using electron microscopy and flow cytometry.RNA-interference-mediated knockdown of Phb1(PHBi)was performed in BRL-3A cells.RESULTS Compared with sham-operation control groups,Phb1 mRNA and protein levels in 70%PHx test groups were downregulated at 24 h,then upregulated at 72 and 168 h.Phb1 was mainly located in mitochondria,showed a reduced abundance at 24 h,significantly increased at 72 h,and almost recovered to normal at 168 h.Phb1 was also present in nuclei,with continuous increase in abundance observed 72 and 168 h after 70%PHx.The altered ultrastructure and reduced mass of mitochondria during LR had almost completely recovered to normal at 168 h.PHBi in BRL-3A cells resulted in increased S-phase entry,a higher number of apoptotic cells,and disruption of mitochondrial membrane potential.CONCLUSION Phb1 may contribute to maintaining mitochondrial stability and could play a role in regulating cell proliferation and apoptosis of rat liver cells during LR.

Colon signet-ring cell carcinoma with chylous ascites caused by immunosuppressants following liver transplantation:A case report

BACKGROUND Chylous ascites is caused by disruption of the lymphatic system,which is characterized by the accumulation of a turbid fluid containing high levels of triglycerides within the abdominal cavity.The two most common causes are cirrhosis and tuberculosis,and colon signer ring cell carcinoma(SRCC)due to the use of immunosuppressants is extremely rare in cirrhotic patients after liver transplantation,making it prone to misdiagnosis and missed diagnosis.CASE SUMMARY A 52-year-old man who underwent liver transplantation and was administered with immunosuppressants for 8 months was admitted with a 3-month history of progressive abdominal distention.Initially,based on lymphoscintigraphy and lymphangiography,lymphatic obstruction was considered,and cystellar chyli decompression with band lysis and external membrane stripping of the lymphatic duct was performed.However,his abdominal distention was persistent without resolution.Abdominal paracentesis revealed allogenic cells in the ascites,and immunohistochemistry analysis revealed adenocarcinoma cells with phenotypic features suggestive of a gastrointestinal origin.Gastrointestinal endoscopy was performed,and biopsy showed atypical signet ring cells in the ileocecal valve.The patient eventually died after a three-month follow-up due to progression of the tumor.CONCLUSION Colon SRCC,caused by immunosuppressants,is an unusual but un-neglected cause of chylous ascites.

Prognostic role of the stromal cell derived factor-1 in patients with hepatitis B virus-related acute-on-chronic liver failure

BACKGROUND Stromal cell derived factor-1(SDF-1)plays a pivotal role in the recruitment of stem cells to injured livers.However,the changes of SDF-l in patients with hepatitis B virus(HBV)-related acute-on-chronic liver failure(ACLF)have yet to be elucidated.AIM To study the SDF-1 changes in patients with HBV-related ACLF.METHODS 30 patients with HBV-related ACLF,27 patients with chronic hepatitis B and 20 healthy individuals are involved in our study.The SDF-l mRNA expression in liver tissue was detected by quantitative real-time polymerase chain reaction.Immunohistochemical staining was performed to illustrate the expression of SDFl,CXC receptor 4(CXCR4)and Ki67.The serum SDF-l concentrations were also detected by enzyme-linked immunosorbent assays.RESULTS The expression of SDF-1 mRNA from ACLF patients was remarkably higher than that from other patients(both P<0.05).The expression of SDF-l,CXCR4 and Ki67 from ACLF were the highest among the three groups(all P<0.01).The serum SDF-l levels in ACLF patients were significantly lower than that in other patients(both P<0.01).Moreover,in ACLF patients,the serum SDF-1 Levels were positively correlated with serum total bilirubin and international normalized ratio.In addition,the serum SDF-l levels in survival were significantly lower compared with the non-survivals(P<0.05).The area under the curve for the serum SDF-1 level in predicting 28-d mortality was 0.722(P<0.05).CONCLUSION This study provides the SDF-1 changes in patients with HBV-related ACLF.The SDF-1 Level at admission may serve as a promising prognostic marker for predicting short-term prognosis.

Single-cell RNA sequencing reveals the dynamics of hepatic non-parenchymal cells in autoprotection against acetaminophen-induced hepatotoxicity

Gaining a better understanding of autoprotection against drug-induced liver injury(DILI)may provide new strategies for its prevention and therapy.However,little is known about the underlying mechanisms of this phenomenon.We used single-cell RNA sequencing to characterize the dynamics and functions of hepatic non-parenchymal cells(NPCs)in autoprotection against DILI,using acetaminophen(APAP)as a model drug.Autoprotection was modeled through pretreatment with a mildly hepatotoxic dose of APAP in mice,followed by a higher dose in a secondary challenge.NPC subsets and dynamic changes were identified in the APAP(hepatotoxicity-sensitive)and APAP-resistant(hepatotoxicity-resistant)groups.A chemokine(C-C motif)ligand 2^(+)endothelial cell subset almost disappeared in the APAP-resistant group,and an R-spondin 3^(+)endothelial cell subset promoted hepatocyte proliferation and played an important role in APAP autoprotection.Moreover,the dendritic cell subset DC-3 may protect the liver from APAP hepatotoxicity by inducing low reactivity and suppressing the autoimmune response and occurrence of inflammation.DC-3 cells also promoted angiogenesis through crosstalk with endothelial cells via vascular endothelial growth factor-associated ligand-receptor pairs and facilitated liver tissue repair in the APAP-resistant group.In addition,the natural killer cell subsets NK-3 and NK-4 and the Sca-1^(-)CD62L^(+)natural killer T cell subset may promote autoprotection through interferon-γ-dependent pathways.Furthermore,macrophage and neutrophil subpopulations with anti-inflammatory phenotypes promoted tolerance to APAP hepatotoxicity.Overall,this study reveals the dynamics of NPCs in the resistance to APAP hepatotoxicity and provides novel insights into the mechanism of autoprotection against DILI at a high resolution.

Hepatic non-parenchymal cells and extracellular matrix participate in oval cell-mediated liver regeneration

AIM： To elucidate the interaction between non- parenchymal cells, extracellular matrix and oval cells during the restituting process of liver injury induced by partial hepatectomy （PH）. METHODS： We examined the localization of oval cells, non-parenchymal cells, and the extracellular matrix components using immunohistochemical and double immunofluorescent analysis during the proliferation and differentiation of oval cells in N-2- acetylaminofluorene （2-AAF）/PH rat model. RESULTS： By day 2 after PH, small oval cells began to proliferate around the portal area. Most of stellate cells and laminin were present along the hepatic sinusoids in the periportal area. Kupffer cells and fibronectin markedly increased in the whole hepatic Iobule. From day 4 to 9, oval cells spread further into hepatic parenchyma, closely associated with stellate cells, fibronectin and laminin. Kupffer cells admixed with oval cells by day 6 and then decreased in the periportal zone. From day 12 to 15, most of hepatic stellate cells （HSCs）, laminin and fibronectin located around the small hepatocyte nodus, and minority of them appeared in the nodus. Kupffer cells were mainly limited in the pericentral sinusoids. After day 18, the normal liver Iobule structures began to recover.CONCLUSION： Local hepatic microenvironment may participate in the oval cell-mediated liver regeneration through the cell-cell and cell-matrix interactions.

Single-cell transcriptomic dissection of the cellular and molecular events underlying the triclosan-induced liver fibrosis in mice

Background: Triclosan [5-chloro-2-(2,4-dichlorophenoxy) phenol, TCS], a common antimicrobial additive in many personal care and health care products, is frequently detected in human blood and urine. Therefore, it has been considered an emerging and potentially toxic pollutant in recent years. Long-term exposure to TCS has been suggested to exert endocrine disruption effects, and promote liver fibrogenesis and tumorigenesis. This study was aimed at clarifying the underlying cellular and molecular mechanisms of hepatotoxicity effect of TCS at the initiation stage.Methods: C57BL/6 mice were exposed to different dosages of TCS for 2 weeks and the organ toxicity was evaluated by various measurements including complete blood count, histological analysis and TCS quantification. Single cell RNA sequencing(scRNA-seq) was then carried out on TCS-or mock-treated mice livers to delineate the TCS-induced hepatotoxicity. The acquired single-cell transcriptomic data were analyzed from different aspects including differential gene expression, transcription factor(TF) regulatory network, pseudotime trajectory, and cellular communication, to systematically dissect the cellular and molecular events after TCS exposure. To verify the TCS-induced liver fibrosis,the expression levels of key fibrogenic proteins were examined by Western blotting, immunofluorescence, Masson’s trichrome and Sirius red stainings. In addition, normal hepatocyte cell MIHA and hepatic stellate cell LX-2 were used as in vitro cell models to experimentally validate the effects of TCS by immunological, proteomic and metabolomic technologies.Results: We established a relatively short term TCS exposure murine model and found the TCS mainly accumulated in the liver. The scRNA-seq performed on the livers of the TCS-treated and control groups profiled the gene expressions of > 76,000 cells belonging to 13 major cell types. Among these types, hepatocytes and hepatic stellate cells(HSCs)were significantly increased in TCS-treated group. We found that TCS promoted fibrosis-associated proliferation of hepatocytes, in which Gata2 and Mef2c are the key driving TFs. Our data also suggested that TCS induced the proliferation and activation of HSCs, which was experimentally verified in both liver tissue and cell model. In addition,other changes including the dysfunction and capillarization of endothelial cells, an increase of fibrotic characteristics in B plasma cells, and M2 phenotype-skewing of macrophage cells, were also deduced from the scRNA-seq analysis, and these changes are likely to contribute to the progression of liver fibrosis. Lastly, the key differential ligand-receptor pairs involved in cellular communications were identified and we confirmed the role of GAS6_AXL interactionmediated cellular communication in promoting liver fibrosis.Conclusions: TCS modulates the cellular activities and fates of several specific cell types(including hepatocytes, HSCs,endothelial cells, B cells, Kupffer cells and liver capsular macrophages) in the liver, and regulates the ligand-receptor interactions between these cells, thereby promoting the proliferation and activation of HSCs, leading to liver fibrosis.Overall, we provide the first comprehensive single-cell atlas of mice livers in response to TCS and delineate the key cellular and molecular processes involved in TCS-induced hepatotoxicity and fibrosis.

Angiotensin-converting enzyme 2 improves liver fibrosis in mice by regulating autophagy of hepatic stellate cells

BACKGROUND Liver fibrosis is the common pathological process associated with the occurrence and development of various chronic liver diseases.At present,there is still a lack of effective prevention and treatment methods in clinical practice.Hepatic stellate cell(HSC)plays a key role in liver fibrogenesis.In recent years,the study of liver fibrosis targeting HSC autophagy has become a hot spot in this research field.Angiotensin-converting enzyme 2(ACE2)is a key negative regulator of reninangiotensin system,and its specific molecular mechanism on autophagy and liver fibrosis needs to be further explored.AIM To investigate the effect of ACE2 on hepatic fibrosis in mice by regulating HSC autophagy through the Adenosine monophosphate activates protein kinases(AMPK)/mammalian target of rapamycin(mTOR)pathway.METHODS Overexpression of ACE2 in a mouse liver fibrosis model was induced by injection of liver-specific recombinant adeno-associated virus ACE2 vector(rAAV2/8-ACE2).The degree of liver fibrosis was assessed by histopathological staining and the biomarkers in mouse serum were measured by Luminex multifactor analysis.The number of apoptotic HSCs was assessed by terminal deoxynucleoitidyl transferase-mediated dUTP nick-end labeling(TUNEL)and immunofluorescence staining.Transmission electron microscopy was used to identify the changes in the number of HSC autophagosomes.The effect of ACE2 overexpression on Wu Y et al.ACE2 improves liver fibrosis through autophagy WJG https://www.wjgnet.com 4976 September 7,2023 Volume 29 Issue 33 autophagy-related proteins was evaluated by multicolor immunofluorescence staining.The expression of autophagy-related indicators and AMPK pathway-related proteins was measured by western blotting.RESULTS A mouse model of liver fibrosis was successfully established after 8 wk of intraperitoneal injection of carbon tetrachloride(CCl4).rAAV2/8-ACE2 administration reduced collagen deposition and alleviated the degree of liver fibrosis in mice.The serum levels of platelet-derived growth factor,angiopoietin-2,vascular endothelial growth factor and angiotensin II were decreased,while the levels of interleukin(IL)-10 and angiotensin-(1-7)were increased in the rAAV2/8-ACE2 group.In addition,the expression of alpha-smooth muscle actin,fibronectin,and CD31 was down-regulated in the rAAV2/8-ACE2 group.TUNEL and immunofluorescence staining showed that rAAV2/8-ACE2 injection increased HSC apoptosis.Moreover,rAAV2/8-ACE2 injection notably decreased the number of autophagosomes and the expression of autophagy-related proteins(LC3I,LC3II,Beclin-1),and affected the expression of AMPK pathway-related proteins(AMPK,p-AMPK,p-mTOR).CONCLUSION ACE2 overexpression can inhibit HSC activation and promote cell apoptosis by regulating HSC autophagy through the AMPK/mTOR pathway,thereby alleviating liver fibrosis and hepatic sinusoidal remodeling.